Primary cSCCs Research Articles

Cancer-associated fibroblast activation predicts progression, metastasis, and prognosis of cutaneous squamous cell carcinoma.

Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and the metastatic disease is associated with poor prognosis. Cancer-associated fibroblasts (CAFs) promote progression of cancer, but their role in cSCC is largely unknown. We examined the potential of CAF markers in the assessment of metastasis risk and prognosis of primary cSCC. We utilized multiplexed fluorescence immunohistochemistry for profiling CAF landscape in metastatic and non-metastatic primary human cSCCs, in metastases, and in premalignant epidermal lesions. Quantitative high-resolution image analysis was performed with two separate panels of antibodies for CAF markers and results were correlated with clinical and histopathological parameters including disease-specific mortality. Increased stromal expression of fibroblast activation protein (FAP), α-smooth muscle actin, and secreted protein acidic and rich in cysteine (SPARC) were associated with progression to invasive cSCC. Elevation of FAP and platelet-derived growth factor receptor-β (PDGFRβ) expression was associated with metastasis risk of primary cSCCs. High expression of PDGFRβ and periostin correlated with poor prognosis. Multimarker combination defined CAF subset, PDGFRα-/PDGFRβ+/FAP+, was associated with invasion and metastasis, and independently predicted poor disease-specific survival. These results identify high PDGFRβ expression alone and multimarker combination PDGFRα-/PDGFRβ+/FAP+ by CAFs as potential biomarkers for risk of metastasis and poor prognosis.

Abstract LB207: C5ar1 is a novel biomarker and potential therapeutic target in advanced cutaneous squamous cell carcinoma

Abstract Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and the metastatic disease is associated with poor prognosis. At present, there are no established molecular markers to predict the metastasis risk and prognosis of patients with primary cSCC. Recently, the role of specific complement components in the progression of cSCC has been shown. Here, we have examined the role of complement C5a receptor C5aR1 in the progression and metastasis of cSCC. The results show that the expression of C5aR1 is increased in cSCC cells in 3D spheroid coculture model in the presence of fibroblasts and that treatment with recombinant C5a increases invasion of cSCC cells in this model. Staining for C5aR1 was detected on the surface of tumor cells in the invasive edge of human cSCC xenograft tumors in vivo. Staining of metastatic and non-metastatic primary human cSCCs, premalignant epidermal lesions and normal skin for C5aR1 with multiplexed fluorescence and chromogenic immunohistochemistry revealed stronger expression of C5aR1 on surface of cSCC tumor cells compared to epidermal cells in cSCC in situ, actinic keratosis and normal skin. Increased expression of C5aR1 on tumor cell surface was associated with the metastasis risk of primary cSCCs and independently predicted poor disease-specific survival of patients with primary cSCC. These results reveal the role of C5aR1 in cSCC progression and identify C5aR1 as a novel biomarker for metastasis risk and poor prognosis of patients with cSCC. The results also suggest C5aR1 as a novel therapeutic target for treatment of locally advanced and metastatic cSCC. Citation Format: Lauri Heiskanen, Liisa Nissinen, Elina Siljamäki, Jaakko Knuutila, Teijo Pellinen, Markku Kallajoki, Jyrki Heino, Pilvi Riihilä, Veli-Matti Kähäri. C5ar1 is a novel biomarker and potential therapeutic target in advanced cutaneous squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB207.

The combined immunohistochemical expression of AMBRA1 and SQSTM1 identifies patients with poorly differentiated cutaneous squamous cell carcinoma at risk of metastasis: A proof of concept study.

Cutaneous squamous cell carcinoma (cSCC) incidence continues to increase globally with, as of yet, an unmet need for reliable prognostic biomarkers to identify patients at increased risk of metastasis. The aim of the present study was to test the prognostic potential of the combined immunohistochemical expression of the autophagy regulatory biomarkers, AMBRA1 and SQSTM1, to identify high-risk patient subsets. A retrospective cohort of 68 formalin-fixed paraffin-embedded primary cSCCs with known 5-year metastatic outcomes were subjected to automated immunohistochemical staining for AMBRA1 and SQSTM1. Digital images of stained slides were annotated to define four regions of interest: the normal and peritumoral epidermis, the tumor mass, and the tumor growth front. H-score analysis was used to semi-quantify AMBRA1 or SQSTM1 expression in each region of interest using Aperio ImageScope software, with receiver operator characteristics and Kaplan-Meier analysis used to assess prognostic potential. The combined loss of expression of AMBRA1 in the tumor growth front and SQSTM1 in the peritumoral epidermis identified patients with poorly differentiated cSCCs at risk of metastasis (*p < 0.05). Collectively, these proof of concept data suggest loss of the combined expression of AMBRA1 in the cSCC growth front and SQSTM1 in the peritumoral epidermis as a putative prognostic biomarker for poorly differentiated cSCC.

Cumulative incidence and risk factors for cutaneous squamous cell carcinoma metastases in organ transplant recipients: The Skin Care in Organ Transplant Patients in Europe-International Transplant Skin Cancer Collaborative metastases study, a prospective multicenter study

Solid organ transplant recipients (SOTRs) are believed to have an increased risk of metastatic cutaneous squamous cell carcinoma (cSCC), but reliable data are lacking regarding the precise incidence and associated risk factors. In a prospective cohort study, including 19 specialist dermatology outpatient clinics in 15 countries, patient and tumor characteristics were collected using standardized questionnaires when SOTRs presented with a new cSCC. After a minimum of 2years of follow-up, relevant data for all SOTRs were collected. Cumulative incidence of metastases was calculated by the Aalen-Johansen estimator. Fine and Gray models were used to assess multiple risk factors for metastases. Of 514 SOTRs who presented with 623 primary cSCCs, metastases developed in 37 with a 2-year patient-based cumulative incidence of 6.2%. Risk factors for metastases included location in the head and neck area, local recurrence, size > 2cm, clinical ulceration, poor differentiation grade, perineural invasion, and deep invasion. A high-stage tumor that is also ulcerated showed the highest risk of metastasis, with a 2-year cumulative incidence of 46.2% (31.9%-68.4%). SOTRs have a high risk of cSCC metastases and well-established clinical and histologic risk factors have been confirmed. High-stage, ulcerated cSCCs have the highest risk of metastasis.

Personalised decision making to predict absolute metastatic risk in cutaneous squamous cell carcinoma: development and validation of a clinico-pathological model

Cutaneous squamous cell carcinoma (cSCC) is a common skin cancer, affecting more than 2 million people worldwide yearly and metastasising in 2-5% of patients. However, current clinical staging systems do not provide estimates of absolute metastatic risk, hence missing the opportunity for more personalised treatment advice. We aimed to develop a clinico-pathological model that predicts the probability of metastasis in patients with cSCC. Nationwide cohorts from (1) all patients with a first primary cSCC in The Netherlands in 2007-2008 and (2) all patients with a cSCC in 2013-2015 in England were used to derive nested case-control cohorts. Pathology records of primary cSCCs that originated a loco-regional or distant metastasis were identified, and these cSCCs were matched to primary cSCCs of controls without metastasis (1:1 ratio). The model was developed on the Dutch cohort (n=390) using a weighted Cox regression model with backward selection and validated on the English cohort (n=696). Model performance was assessed using weighted versions of the C-index, calibration metrics, and decision curve analysis; and compared to the Brigham and Women's Hospital (BWH) and the American Joint Committee on Cancer (AJCC) staging systems. Members of the multidisciplinary Skin Cancer Outcomes (SCOUT) consortium were surveyed to interpret metastatic risk cutoffs in a clinical context. Eight out of eleven clinico-pathological variables were selected. The model showed good discriminative ability, with an optimism-corrected C-index of 0.80 (95% Confidence interval (CI) 0.75-0.85) in the development cohort and a C-index of 0.84 (95% CI 0.81-0.87) in the validation cohort. Model predictions were well-calibrated: the calibration slope was 0.96 (95% CI 0.76-1.16) in the validation cohort. Decision curve analysis showed improved net benefit compared to current staging systems, particularly for thresholds relevant for decisions on follow-up and adjuvant treatment. The model is available as an online web-based calculator (https://emc-dermatology.shinyapps.io/cscc-abs-met-risk/). This validated model assigns personalised metastatic risk predictions to patients with cSCC, using routinely reported histological and patient-specific risk factors. The model can empower clinicians and healthcare systems in identifying patients with high-risk cSCC and offering personalised care/treatment and follow-up. Use of the model for clinical decision-making in different patient populations must be further investigated. PPP Allowance made available by Health-Holland, Top Sector Life Sciences & Health, to stimulate public-private partnerships.

Cutaneous squamous cell carcinoma-derived extracellular vesicles exert an oncogenic role by activating cancer-associated fibroblasts

Cutaneous squamous cell carcinoma (cSCC) is a fast-increasing cancer with metastatic potential. Extracellular vesicles (EVs) are small membrane-bound vesicles that play important roles in intercellular communication, particularly in the tumor microenvironment (TME). Here we report that cSCC cells secrete an increased number of EVs relative to normal human epidermal keratinocytes (NHEKs) and that interfering with the capacity of cSCC to secrete EVs inhibits tumor growth in vivo in a xenograft model of human cSCC. Transcriptome analysis of tumor xenografts by RNA-sequencing enabling the simultaneous quantification of both the human and the mouse transcripts revealed that impaired EV-production of cSCC cells prominently altered the phenotype of stromal cells, in particular genes related to extracellular matrix (ECM)-formation and epithelial-mesenchymal transition (EMT). In line with these results, co-culturing of human dermal fibroblasts (HDFs) with cSCC cells, but not with normal keratinocytes in vitro resulted in acquisition of cancer-associated fibroblast (CAF) phenotype. Interestingly, EVs derived from metastatic cSCC cells, but not primary cSCCs or NHEKs, were efficient in converting HDFs to CAFs. Multiplex bead-based flow cytometry assay and mass-spectrometry (MS)-based proteomic analyses revealed the heterogenous cargo of cSCC-derived EVs and that especially EVs derived from metastatic cSCCs carry proteins associated with EV-biogenesis, EMT, and cell migration. Mechanistically, EVs from metastatic cSCC cells result in the activation of TGFβ signaling in HDFs. Altogether, our study suggests that cSCC-derived EVs mediate cancer-stroma communication, in particular the conversion of fibroblasts to CAFs, which eventually contribute to cSCC progression.

P26 Risk factors for complications in cutaneous squamous cell carcinoma and proposal of a simplified follow-up strategy, making use of available healthcare resources

Abstract Cutaneous squamous cell carcinoma (cSCC) is a common malignancy, generally cured by surgical excision. A minority of such tumours recur, metastasize or even cause death. The 2021 British Association of Dermatologists (BAD) guidelines outline high-risk features that are used to guide the follow-up of patients after a primary cSCC. The aims of this study were to identify histological features of cSCC that are significantly associated with the development of complications, and to compare the sensitivity, specificity and number of appointments required to follow-up patients using different follow-up criteria. In total, 556 cases of primary invasive cSCCs excised between January 2014 and March 2018 were included and followed-up for a minimum of 4 years. Five per cent (n = 26) of tumours were associated with at least one cSCC-related complication, namely, local recurrence 3.8% (n = 21), nodal metastasis 1.6% (n = 9), distant metastasis 0.4% (n = 2) and disease-specific death 0.7% (n = 4). The initial diagnosed complication was local recurrence in 20 of 26 cases. Only two patients had the first complication beyond 2 years of the original cSCC. Factors that were statistically significantly associated with the development of a complication (P &amp;lt; 0.001) are larger diameter, larger depth of invasion, tumour invasion beyond the dermis, perineural invasion (irrespective of size), lymphovascular invasion, gross bone erosion and deep margin &amp;lt; 1 mm. Differentiation and peripheral margin were not significantly associated with complications. Therefore, it may be proposed that patients with tumours with at least one of the following risk factors be followed-up: diameter &amp;gt; 40 mm; depth of invasion &amp;gt; 6 mm; invasion beyond the dermis; presence of perineural or lymphovascular invasion or deep margin &amp;lt; 1 mm. In adopting these criteria, 739 appointments in 156 patients would be required over 2 years (4 monthly during the first year and 6 monthly during the second year). Twenty of the patients who developed a complication would be followed-up (sensitivity 76.9%, specificity 69.4%). Using the BAD guidelines, up to four more patients with complications would have been followed-up (sensitivity 92.3%, specificity 44.7%), but this would have required a total of 1556 follow-up visits in 242 patients. Given the limited resources, patient burden and the fact that local recurrence is most often the first complication, it may be reasonable to offer routine follow-up only to patients who have at least one of the proposed criteria. Coupled with patient education, this strategy could greatly reduce the number of follow-up visits required. Such a strategy may allow resources to be utilized to reduce waiting times, particularly for 2-week wait referrals, decreasing the delay in new skin cancer diagnoses.

DS08 (P26) Risk factors for complications in cutaneous squamous cell carcinoma and proposal of a simplified follow-up strategy: making use of available healthcare resources

Abstract Cutaneous squamous cell carcinoma (cSCC) is a common malignancy, generally cured by surgical excision. A minority of such tumours recur, metastasize or even cause death. The 2021 British Association of Dermatologists (BAD) guidelines outline high-risk features that are used to guide the follow-up of patients after a primary cSCC. The aims of this study were to identify histological features of cSCC that are significantly associated with the development of complications and to compare the sensitivity, specificity and number of appointments required to follow-up patients using different follow-up criteria. In total, 556 cases of primary invasive cSCCs excised between January 2014 and March 2018 were included and followed-up for a minimum of 4 years. Five per cent (n = 26) of tumours were associated with at least one cSCC-related complication, namely, local recurrence 3.8% (n = 21), nodal metastasis 1.6% (n = 9), distant metastasis 0.4% (n = 2) and disease-specific death 0.7% (n = 4). The initial diagnosed complication was local recurrence in 20 of 26 cases. Only two patients had the first complication beyond 2 years of the original cSCC. Factors that were statistically significantly associated with the development of a complication (P &amp;lt; 0.001) were larger diameter, larger depth of invasion, tumour invasion beyond the dermis, perineural invasion (irrespective of size), lymphovascular invasion, gross bone erosion and deep margin &amp;lt; 1 mm. Differentiation and peripheral margin were not significantly associated with complications. Therefore, it may be proposed that patients with tumours with at least one of the following risk factors be followed-up: diameter &amp;gt; 40 mm, depth of invasion &amp;gt; 6 mm, invasion beyond the dermis, presence of perineural or lymphovascular invasion or deep margin &amp;lt; 1 mm. In adopting these criteria, 739 appointments in 156 patients would be required over 2 years (4-monthly during the first year and 6-monthly during the second year). Twenty of the patients who developed a complication would be followed-up (sensitivity 76.9%, specificity 69.4%). Using the BAD guidelines, up to four more patients with complications would have been followed-up (sensitivity 92.3%, specificity 44.7%), but this would have required a total of 1556 follow-up visits in 242 patients. Given the limited resources, patient burden and the fact that local recurrence is most often the first complication, it may be reasonable to offer routine follow-up only to patients who have at least one of the proposed criteria. Coupled with patient education, this strategy could greatly reduce the number of follow-up visits required. Such a strategy may allow resources to be utilized to reduce waiting times, particularly for 2-week wait referrals, decreasing the delay in new skin cancer diagnoses.

010 Enhanced outcome prediction in cutaneous squamous cell carcinoma using deep-learning and computational histopathology (cSCCnet)

Abstract Cutaneous squamous cell carcinoma (cSCC) is a common skin cancer and leads to a significant disease burden. Although metastases occur in &amp;lt; 5% of patients, effective treatments are limited and 5-year survival is poor, with mortality rates equivalent to those of melanoma in some regions of the world. Despite recent advances in understanding the molecular pathogenesis of cSCC, patient risk stratification is limited and based on clinicopathological staging, with few validated prognostic biomarkers available to guide management [de Jong E, Lammerts MUPA, Genders RE, Bowes Bavinck JN. Update of advanced cutaneous squamous cell carcinoma. J Eur Acad Dermatol Venereol 2022; 36(Suppl. 1):6–10]. In this study, we aimed to use artificial intelligence technology to develop a deep learning model using digitized whole-slide images of cSCC to quantify the risk of metastasis. Routine haematoxylin and eosin-stained slides of 256 primary cSCCs from immunocompetent patients at four collaborating centres were scanned. The area of interest encompassing tumour and peritumoral stroma was annotated, and 512 × 512 pixel tiles were generated from each slide. High-quality tiles (containing sufficient tumour content) were pooled together, with patient-level labels inherited by tiles. We trained multiple models using an 80:10:10 split for training, validation and testing, and the model achieving the highest tile- and slide-level validation accuracy was identified (ResNet50). The final model, namely, cSCCnet, achieved 88% sensitivity, 73% specificity, 64% positive predictive value and 92% negative predictive values on the independent test set. Blinded review of the top predictive tiles by three histopathologists suggested that the predictive features identified by cSCCnet were distinct from histopathological features commonly used by clinicians to estimate metastatic risk. We have demonstrated that it is possible to train a deep learning model using digital pathology to predict the risk of metastasis. cSCCnet surpassed the published performance of currently used staging systems, as well as commercially available gene panel testing (de Jong et al.; Wysong A, Newman JG, Covington KR et al. Validation of a 40-gene expression profile test to predict metastatic risk in localized high-risk cutaneous squamous cell carcinoma. J Am Acad Dermatol 2021; 84:361–9). The morphological features and patterns identified by cSCCnet are different from established histopathological high-risk criteria, indicating that this technology can enhance current practice. Although further validation in a prospective cohort is pending, this is an important step toward creating a risk stratification tool that can be incorporated into routine pathology workflows, significantly improving patient outcomes for this common cancer.

Abstract 2101: Cancer-associated fibroblast activation predicts progression, metastasis, and prognosis of cutaneous squamous cell carcinoma

Abstract Purpose: Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and the metastatic disease is associated with poor prognosis. At present, no established biomarkers are available for predicting the metastasis risk and prognosis of patients with cSCC. Cancer-associated fibroblasts (CAFs) promote the progression of cancer, but their role in cSCC is largely unknown. Here, we have examined the potential of CAF markers in the assessment of metastasis risk and prognosis of primary cSCC. Experimental Design: We utilized multiplexed fluorescence immunohistochemistry for profiling of CAF landscape in metastatic and non-metastatic primary human cSCCs, the actual metastases, and in premalignant and benign epidermal lesions. Quantitative high-resolution image analysis was performed with two separate panels of antibodies for CAF markers and the results were correlated with clinical and histopathological parameters including disease-specific mortality. Results: Increased stromal expression of fibroblast activation protein (FAP), α-smooth muscle actin, and secreted protein acidic and rich in cysteine (SPARC) were associated with progression of premalignant epidermal lesions to invasive cSCC. Furthermore, elevation of FAP and platelet-derived growth factor receptor-β (PDGFRβ) expression was associated with the risk of metastasis of primary cSCCs. High expression of PDGFRβ and periostin correlated with poor prognosis. Furthermore, a multi-marker defined CAF subset, PDGFRα-/PDGFRβ+/FAP+, was associated with invasion and metastasis and independently predicted poor disease-specific survival. Conclusions: These results show, that fibroblast activation evolves during the progression of cSCC. High PDGFRβ expression alone and multimarker combination PDGFRα-/PDGFRβ+/FAP+ are potential biomarkers for the progression of primary cSCC to metastatic stage, and for poor prognosis. Citation Format: Jaakko S. Knuutila, Pilvi Riihilä, Liisa Nissinen, Roosa Kallionpää, Teijo Pellinen, Veli-Matti Kähäri. Cancer-associated fibroblast activation predicts progression, metastasis, and prognosis of cutaneous squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2101.

Loss of dyskerin facilitates the acquisition of metastatic traits by altering the mevalonate pathway.

The initial dissemination of cancer cells from many primary tumors implies intravasation to lymphatic nodes or blood vessels. To investigate the mechanisms involved, we analyzed the expression of small non-coding RNAs in cutaneous squamous cell carcinoma (cSCC), a prevalent tumor that mainly spreads to lymph nodes. We report the reduced expression of small nucleolar RNAs in primary cSCCs that metastasized when compared to non-metastasizing cSCCs, and the progressive loss of DKC1 (dyskerin, which stabilizes the small nucleolar RNAs) along the metastasis. DKC1 depletion in cSCC cells triggered lipid metabolism by altering the mevalonate pathway and the acquisition of metastatic traits. Treatment of DKC1-depleted cells with simvastatin, an inhibitor of the mevalonate pathway, blocked the expression of proteins involved in the epithelial-to-mesenchymal transition. Consistently, the expression of the enzyme 3-hydroxy-3-methylglutaryl-CoA synthase 1 was associated with pathological features of high metastatic risk in cSCC patients. Our data underpin the relevance of the mevalonate metabolism in metastatic dissemination and pave the possible incorporation of therapeutic approaches among the antineoplastic drugs used in routine patient care.

HLA expression as a risk factor for metastases of cutaneous squamous-cell carcinoma in organ- transplant recipients

BackgroundSolid organ-transplant recipients (SOTR) have an increased risk of cutaneous squamous-cell carcinoma (cSCC), metastasis and death from cSCC. In immunocompetent patients with mucosal SCC, downregulation of HLA class I is associated with poor prognosis. Since the degree of HLA expression on tumor cells could play a role in immunogenicity and pathophysiology of cSCC metastasis, we hypothesized that decreased HLA expression is associated with an increased risk of metastasis. MethodsWe compared HLA expression between primary metastasized cSCCs, their metastases, and non-metastasized cSCCs from the same patients. Samples were stained for HLA-A, HLA-B/-C and quantified by calculating the difference in immunoreactivity score (IRS) of the primary cSCC compared with all non-metastasized cSCCs. ResultsThe mean IRS score for HLA-B/C expression was 2.07 point higher in metastasized compared to non-metastasized cSCCs (p = 0.065, 95 % CI −0.18–4.32). 83.3 % of the primary metastasized cSCCs had an IRS score of 4 or higher, compared to 42.9 % in non-metastasized cSCCs. Moderately to poorly differentiated cSCCs had more HLA class I expression compared to well-differentiated cSCCs. ConclusionContrary to immunocompetent patients, HLA-B/C expression tends to be upregulated in metastasized cSCC compared to non-metastasized cSCC in SOTR, suggesting that different tumor escape mechanisms play a role in SOTR compared to immunocompetent patients.

466 Cancer-associated fibroblast activation predicts progression, metastasis and prognosis of cutaneous squamous cell carcinoma

Cutaneous squamous cell carcinoma (cSCC) possesses metastatic potential and in metastatic case the prognosis is poor. At present, there are no established biomarkers for assessment of metastasis risk or prognosis of primary cSCC. Cancer-associated fibroblasts (CAFs) promote the progression of cancer, but there is limited evidence of their role in cSCC. Here, we have examined the potential of CAF markers in the assessment of metastasis risk and prognosis of cSCC. We used multiplexed fluorescence immunohistochemistry to profile CAF landscape in metastatic and non-metastatic primary human cSCCs, actual metastases, and in premalignant and benign epidermal lesions.

LB886 Cancer-associated fibroblast activation predicts progression, metastasis and prognosis of cutaneous squamous cell carcinoma

Cutaneous squamous cell carcinoma (cSCC) possesses metastatic potential and in metastatic cases the prognosis is poor. At present, there are no established metastasis risk-associated or prognostic biomarkers for cSCC. Cancer-associated fibroblasts (CAFs) promote the progression of cancer, but there is limited evidence for their role in cSCC. We have examined the potential of CAF-associated biomarkers in the assessment of metastasis risk and prognosis of cSCC. We used multiplex immunohistochemistry to profile CAF landscape in tissue microarrays containing metastatic (n=33) and non-metastatic (n=146) primary human cSCCs, metastases (n=13), cSCC in situ (cSCCIS) (n=59), actinic keratoses (AK) (n=67), seborrheic keratoses (n=17) and normal skin (n=73).

Clinical outcomes of high-risk cutaneous squamous cell carcinomas treated with Mohs surgery alone: An analysis of local recurrence, regional nodal metastases, progression-free survival, and disease-specific death

BackgroundThe incidence of cutaneous squamous cell carcinoma (cSCC) continues to increase, and it is now predicted that the number of deaths from cSCC will surpass that of melanoma within the next 5 years. Although most cSCCs are successfully treated, there exists an important subset of high-risk tumors that have the highest propensity for local recurrence (LR), nodal metastasis (NM), and disease-specific death (DSD). ObjectiveWe investigated the clinical outcomes of high-risk cSCCs treated with Mohs surgery (MS) alone, analyzing LR, NM, distant metastasis, and DSD. In addition, we analyzed progression-free survival and DSD in patients who underwent salvage head/neck dissection for regional NMs. MethodsRetrospective review of all high-risk cSCC treated in our clinics between January 1, 2000, and January 1, 2020, with follow-up through April 1, 2020. SettingTwo university-affiliated, private-practice MS referral centers. ResultsIn total, 581 high-risk primary cSCCs were identified in 527 patients, of which follow-up data were obtained for 579 tumors. The 5-year disease-specific survival was 95.7%, with a mean survival time of 18.6 years. The 5-year LR-free survival was 96.9%, the regional NM-free survival was 93.8%, and the distant metastasis–free survival was 97.3%. The 5- and 10-year progression-free survival rates from metastatic disease were 92.6 and 90.0%, respectively. In patients who experienced regional NMs and underwent salvage head and neck dissection with or without radiation, the 2-year disease-specific survival was 90.5%. ConclusionOur cohort, which is the largest high-risk cSCC cohort treated with MS to date, experienced lower rates of LR, NM, and DSD than those reported with historical reference controls using both the Brigham and Women’s Hospital and American Joint Committee on Cancer, Eighth Edition, staging systems. We demonstrated that MS confers a disease-specific survival advantage over historical wide local excision for high-risk tumors. Moreover, by improving local tumor control, MS appears to reduce the frequency of regional metastatic disease and may confer a survival advantage even for patients who develop regional metastases.

Identification of metastatic primary cutaneous squamous cell carcinoma utilizing artificial intelligence analysis of whole slide images

Cutaneous squamous cell carcinoma (cSCC) harbors metastatic potential and causes mortality. However, clinical assessment of metastasis risk is challenging. We approached this challenge by harnessing artificial intelligence (AI) algorithm to identify metastatic primary cSCCs. Residual neural network-architectures were trained with cross-validation to identify metastatic tumors on clinician annotated, hematoxylin and eosin-stained whole slide images representing primary non-metastatic and metastatic cSCCs (n = 104). Metastatic primary tumors were divided into two subgroups, which metastasize rapidly (≤ 180 days) (n = 22) or slowly (> 180 days) (n = 23) after primary tumor detection. Final model was able to predict whether primary tumor was non-metastatic or rapidly metastatic with slide-level area under the receiver operating characteristic curve (AUROC) of 0.747. Furthermore, risk factor (RF) model including prediction by AI, Clark’s level and tumor diameter provided higher AUROC (0.917) than other RF models and predicted high 5-year disease specific survival (DSS) for patients with cSCC with 0 or 1 RFs (100% and 95.7%) and poor DSS for patients with cSCCs with 2 or 3 RFs (41.7% and 40.0%). These results indicate, that AI recognizes unknown morphological features associated with metastasis and may provide added value to clinical assessment of metastasis risk and prognosis of primary cSCC.

Matrix Effectors in the Pathogenesis of Keratinocyte-Derived Carcinomas.

Basal cell carcinoma (BCC) and cutaneous squamous cell carcinoma (cSCC), referred to as keratinocyte carcinomas, are skin cancer with the highest incidence. BCCs, rarely metastasize; whereas, though generally not characterized by high lethality, approximately 2–4% of primary cSCCs metastasize with patients exhibiting poor prognosis. The extracellular matrix (ECM) serves as a scaffold that provides structural and biological support to cells in all human tissues. The main components of the ECM, including fibrillar proteins, proteoglycans (PGs), glycosaminoglycans (GAGs), and adhesion proteins such as fibronectin, are secreted by the cells in a tissue-specific manner, critical for the proper function of each organ. The skin compartmentalization to the epidermis and dermis compartments is based on a basement membrane (BM), a highly specialized network of ECM proteins that separate and unify the two compartments. The stiffness and assembly of BM and tensile forces affect tumor progenitors' invasion at the stratified epithelium's stromal border. Likewise, the mechanical properties of the stroma, e.g., stiffness, are directly correlated to the pathogenesis of the keratinocyte carcinomas. Since the ECM is a pool for various growth factors, cytokines, and chemokines, its' intense remodeling in the aberrant cancer tissue milieu affects biological functions, such as angiogenesis, adhesion, proliferation, or cell motility by regulating specific signaling pathways. This review discusses the structural and functional modulations of the keratinocyte carcinoma microenvironment. Furthermore, we debate how ECM remodeling affects the pathogenesis of these skin cancers.

Complement Factor D Is a Novel Biomarker and Putative Therapeutic Target in Cutaneous Squamous Cell Carcinoma.

Simple SummaryThe incidence of the most common metastatic skin malignancy, cutaneous squamous cell carcinoma (cSCC), is growing worldwide, and the prognosis of the metastatic disease is poor. Presently, there are no biomarkers or therapeutic targets for high-risk cSCCs. Recent studies have demonstrated the essential role of autocrine complement synthesis in the progression of cSCC. Here, we have evaluated the role of complement Factor D (FD), the rate-limiting enzyme of the alternative complement pathway, in cSCC development. The results identify FD as a novel biomarker and putative therapeutic target for cSCC and propose the small-molecule FD inhibitor Danicopan as a highly specific drug candidate in the therapy of advanced cSCC. It is expected that the discovery of complement-associated molecular markers for cSCC progression would improve diagnosis, classification, prognostication, and targeted therapy of cSCC and its precursors in the future.Cutaneous squamous cell carcinoma (cSCC) is the most prevalent metastatic skin cancer. Previous studies have demonstrated the autocrine role of complement components in cSCC progression. We have investigated factor D (FD), the key enzyme of the alternative complement pathway, in the development of cSCC. RT-qPCR analysis of cSCC cell lines and normal human epidermal keratinocytes (NHEKs) demonstrated significant up-regulation of FD mRNA in cSCC cells compared to NHEKs. Western blot analysis also showed more abundant FD production by cSCC cell lines. Significantly higher FD mRNA levels were noted in cSCC tumors than in normal skin. Strong tumor cell-associated FD immunolabeling was detected in the invasive margin of human cSCC xenografts. More intense tumor cell-specific immunostaining for FD was seen in the tumor edge in primary and metastatic cSCCs, in metastases, and in recessive dystrophic epidermolysis bullosa-associated cSCCs, compared with cSCC in situ, actinic keratosis and normal skin. FD production by cSCC cells was dependent on p38 mitogen-activated protein kinase activity, and it was induced by interferon-γ and interleukin-1β. Blocking FD activity by Danicopan inhibited activation of extracellular signal-regulated kinase 1/2 and attenuated proliferation of cSCC cells. These results identify FD as a novel putative biomarker and therapeutic target for cSCC progression.

Single-cell transcriptomic analysis reveals the critical molecular pattern of UV-induced cutaneous squamous cell carcinoma

Cutaneous squamous cell carcinoma (cSCC) is the second most common nonmelanoma skin cancer characterized by high invasiveness, heterogeneity, and mainly occurs in the ultraviolet (UV)-exposed regions of the skin, but its pathogenesis is still unclear. Here, we generated single-cell transcriptome profiles for 350 cells from six primary UV-induced cSCCs, together with matched adjacent skin samples, and three healthy control skin tissues by single-cell RNA-sequencing technology based on Smart-seq2 strategy. A series of bioinformatics analyses and in vitro experiments were used to decipher and validate the critical molecular pattern of cSCC. Results showed that cSCC cells and normal keratinocytes were significantly distinct in gene expression and chromosomal copy number variation. Furthermore, cSCC cells exhibited 18 hallmark pathways of cancer by gene set enrichment analysis. Differential expression analysis demonstrated that many members belonging to S100 gene family, SPRR gene family, and FABP5 were significantly upregulated in cSCC cells. Further experiments confirmed their upregulation and showed that S100A9 or FABP5 knockdown in cSCC cells inhibited their proliferation and migration through NF-κB pathway. Taken together, our data provide a valuable resource for deciphering the molecular pattern in UV-induced cSCC at a single-cell level and suggest that S100A9 and FABP5 may provide novel targets for therapeutic intervention of cSCC in the future.

266 Identification of metastatic primary cutaneous squamous cell carcinoma using artificial intelligence analysis of whole slide images

Cutaneous squamous cell carcinoma (cSCC) possesses metastatic potential and causes significant mortality. However, clinical assessment of metastasis risk of primary cSCC is challenging. In this study, we have used artificial intelligence (AI) algorithm to identify primary cSCCs with risk for metastasis. Residual neural network-architectures were trained and fine-tuned on tumor tiles extracted from clinician annotated, hematoxylin and eosin (HE)-stained whole slide images representing primary metastatic (n=45) and non-metastatic (n=59) cSCCs.